What is SDS-PAGE gel? What are reducing and non-reducing conditions?
SDS-PAGE (sodium dodecyl sulfate–polyacrylamide gel electrophoresis) is a method used to separate proteins based on their mass. SDS is an ionic detergent that can denature and bind proteins, imparting them with a uniform negative charge. This means that when an electric current is applied to a polyacrylamide gel, proteins bound to SDS will migrate down the gel towards the positively charged electrode, being separated solely based on size. Proteins can be reduced before running on an SDS-PAGE gel. Under reducing conditions, β-mercaptoethanol (β-ME or 2-ME) or dithiothreitol (DTT) is added, which reduces disulfide bonds in proteins, allowing for better separation by size when run on the gel. Many growth factors are disulfide-linked dimers, and running non-reduced proteins on the same gel as reduced proteins can confirm the correct dimeric state.
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